Antibody Type: Rabbit Polyclonal

Application: WB,IHC-P,IHC-Fr,ICC/IF,IF,FCM

Reactivity: Human,Mouse,Rat (predicted:Chicken,Pig,Cow)

IgG

Human,Mouse,Rat (predicted:Chicken,Pig,Cow)

1. Paraformaldehyde-fixed, paraffin embedded (Mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 min; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (GADD45) Polyclonal Antibody, Unconjugated (TMAB-00733) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.
2. Tissue/cell: rat brain tissue; 4% Paraformaldehyde-fixed and paraffin-embedded;
Antigen retrieval: citrate buffer (0.01M, pH6.0), Boiling bathing for 15 min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30 min; Blocking buffer (normal goat serum) at 37°C for 20 min;
Incubation: Anti-GADD45 Polyclonal Antibody, Unconjugated (TMAB-00733) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody and DAb staining.
3. Sample:
Lane1: Brain (Rat) Lysate at 30 μg
Lane2: Kidney (Rat) Lysate at 30 μg
Primary: Anti-GADD45 (TMAB-00733) at 1:200 dilution;
Secondary: HRP conjugated Goat-Anti-Rabbit Igg (secondary antibody) at 1: 3000 dilution;
Predicted band size: 18 kDa
Observed band size: 18 kDa
We are unsure as to the identity of these extra bands
4. Tissue/cell: A431 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (GADD45) Polyclonal Antibody, Unconjugated (TMAB-00733) 1:200, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (5 μg/ml, blue) was used to stain the cell nucleus.
5. Tissue/cell: U-2OS cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum) at 37°C for 20 min; Antibody incubation with (GADD45) Polyclonal Antibody, Unconjugated (TMAB-00733) 1:200, 90 minutes at 37°C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (5 μg/ml, blue) was used to stain the cell nucleus.
6. Paraformaldehyde-fixed, paraffin embedded (Human liver carcinoma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 min; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (GADD45) Polyclonal Antibody, Unconjugated (TMAB-00733) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.
7. Blank control: A431. Primary Antibody (green line): Rabbit Anti-GADD45 antibody (TMAB-00733)
Dilution: 1 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-AF647
Dilution: 1 μg/test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 0.1% PBST for 20 min at room temperature. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature.

WB,IHC-P,IHC-Fr,ICC/IF,IF,FCM

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; ICC/IF: 1:100-500; IF: 1:100-500; FCM: 3ug/Test

Polyclonal

KLH conjugated synthetic peptide: human GADD45

Human

1647

Base Excision Repair